Summary: A heptavalent pneumococcal polysaccharide-protein conjugate vaccine, Prevnar, and other conjugate vaccines have used tetanus toxoid or diphtheria toxoid as the carrier protein. The use of pneumococcal cell-surface protein antigens, such as pneumolysin, autolysin, or PspA can potentially provide a broader immunity. In addition, they can be used as conjugate carriers for PS antigens to stimulate increased immunity to PSs for protection against pneumococcal infection. Protective immunity against pneumococcal infection is directly correlated with IgG antibody levels, and avidity measurements may provide a better correlation between serum antibody concentration and protective immunity against pneumococcal diseases. It is important to determine the quality, quantity, and duration of these antibodies. The objective of the present study was to evaluate the avidity, serum specific antibodies and their concentrations over time post immunization as well as opsonophagocytic activity of the antibodies. The effect of priming with the 7-valent 9V PS containing conjugate followed by a booster dose of 23-valent PS vaccines was also examined. The relative avidities of IgG and IgM Abs were highest in mice immunized twice with 7-valent conjugate, and combined immunization with 7-valent conjugate followed by 23-valent PS vaccine. Mice that were immunized with 9V PS alone exhibited the lowest avidity. Serum concentrations of 9V PS IgG and IgM Abs were higher in mice immunized with 9V PS-pneumolysin, 9V PS-autolysin conjugates, 7-valent conjugate alone or 7-valent conjugate followed by 23-valent PS vaccine compared to the mice given 9V PS alone. Furthermore, the 9V PS IgG and IgM antibody responses were high in mice immunized with 9V PS-Ply, or 9V PS-Aly conjugate at 1, 4, 8, and 12 weeks after three doses, whereas the antibody responses remained high in mice immunized with 7-valent conjugate twice at 1, 2, and 8 weeks after the final injection. Antisera of mice immunized twice with 7-valent conjugate or with the conjugate followed by 23-valent PS vaccine showed high opsonophagocytic titers. These results indicate that the 9V PS conjugates using pneumococcal proteins, and 7-valent conjugate vaccines exhibit effective immunity against pneumococcal infection, and that priming mice with 7-valent conjugate followed by 23-valent PS vaccine induced high immune responses.